In the fields of medicine, biophysics, biology, pharmacology, material control and image archiving of histological preparations, optical scanning microscopes are required which have a high throughput (that is to say they can optically acquire a large number of objects in a short period of time) and can acquire, in the form of an image, large areas, for example of the order of magnitude of up to 20cm×20cm, with a typical image resolution of approximately 1μm. The intention is further to create depth profiles of the object to be optically scanned for specific applications.
A scanning microscope of this type is usually equipped with an image-producing scanner camera whose pixels are produced in the form of an integrated circuit (chip) in CMOS technology and which scanner camera is arranged on the image plane of the microscope. An objective lens system in the microscope permits different magnifications of object images on the object plane which are typical for microscopy, for example 10-fold, 20-fold, 50-fold, 100-fold, etc.
In order to optically scan an object, a mechanical translation stage moves horizontally under computer control, the two basic directions of this horizontal movement usually being referred to as “x-direction” and as “y-direction”. The movement in the “z-direction”, that is to say the vertical movement and hence the focusing of the microscope, is ensured by means of a computer-controlled linear guidance of the objective lens system. Depending on the magnification and the specifications of the objective lens system, a typical field of view of a microscope of this type has a size of approximately 200μm×200μm and is completely imaged on the image plane on the photosensitive image-recording chip (e.g. CMOS chip) of the scanner camera. Nowadays a color camera having a resolution of more than from 1Mpixel is used as a scanner camera.
For an area of 10cm×10cm to be optically scanned, the scanner camera thus produces about 250,000individual images. This number potentially still needs to be multiplied by the number of the depth profile images. The individual images of each plane, which are recorded in a step-by-step fashion and are located one next to the other, can then be combined by “software stitching” to form one or more total images. The total images can subsequently be analyzed automatically under computer control and depending on the application.
Electronic acquisition of such large amounts of data and the simultaneous image processing and image storing in real time is largely unproblematic and currently achieved at approximately 10-100frames per second (fps) at full image size. Future developments will probably be able to increase this value again by a factor of 10. If an image processing rate of 100fps is taken as a basis, a complete microscopic image acquisition of the area in the abovementioned example of 100cm2is possible electronically in approximately 40minutes. If the intention is for only parts of the area to be acquired microscopically (e.g. an arrangement of object slides or a sample matrix), this number is correspondingly lower.
The mechanical realization of the movement operations, however, poses a problem, since, in order to record an image, the object always has to be brought into a rest position. The mechanics of a commercial mechanical translation stage are, however, not able to carry out such a high number of starting operations (accelerations) and stopping operations (decelerations), in accordance with the above-described example 100starting/stopping operations per second, with simultaneously low travel (ca. 200μm).
Instability of the mechanical system of the scanning microscope occurs additionally on account of inevitable considerable mechanical oscillations which are caused by the starting/stopping operations which occur at short intervals. The number of starting/stopping operations in continuous operation is therefore limited to about 1-3 per second. Necessary longer displacement distances, such as, for example, when relocating specific positions, when moving toward object slides or in the case of multiple samples, are irrelevant in terms of the time balance of a scanning operation.